The optical light microscope has played a most profound role in biological research by permitting the first view into the microscopic world of cells. One of the greatest applications of light microscopy is in the study of living microorganisms and tissues, since the light microscope can probe a sample in its somewhat natural condition. However, the light microscope is limited to a resolution of about one micron (1 micron=10.sup.-4 cm), corresponding to a maximum usable magnification of 2000x. Progress in biology and medicine has demanded must higher magnifications and resolutions than those achievable with traditional light microscopes, and newer technologies in electron microscopy have been developed. A good scanning electron microscope can provide usable magnifications near 200,000x with a resolution in tens of Angstroms (1 A=10.sup.-8 cm).
While optical light microscopes and electron microscopes have contributed to significant scientific discoveries in the past and large numbers of these instruments are currently in use, both devices are of limited usefulness and do not fully address the micro-imaging needs of contemporary research and industry. Optical light microscopes do not provide the magnification power and high resolution required for many industrial applications and current scientific research. Electron microscopes, while providing much higher resolution than traditional optical microscopes, are limited by their inability to image living and wet specimens or other specimens maintained under room conditions.
Alternative imaging technologies, such as computer-aided tomography ("CAT") and nuclear magnetic resonance ("NMR"), allow imaging or living specimens but lack the necessary magnification and high resolution features of electron microscopes.
Generally, electron microscopes operate by directing a beam of electrons rapidly over or through the surface of a specimen contained in an evacuated chamber. As the electrons hit the specimen, the specimen scatters the electrons. A device then reads the dispersion intensities and translates them into an image of the specimen. Obtaining an undistorted image of the specimen from the interaction of the electrons with the specimen requires that the electron beam hit the specimen directly without being scattered by air molecules. To avoid the distortion caused by air molecules, electron microscope specimens are placed in a vacuum chamber during the imaging process, thereby providing the electron beam with an unimpeded path to the specimen.
The vacuum chamber eliminates atmospheric distortion in the imaging process but makes it impossible to use the electron microscope to image living and wet specimens. Living specimens, such as cells and tissues, cannot survive in a vacuum for the time necessary to produce the image. Similarly, wet specimens experience evaporation of their fluid content in the vacuum before an image can be produced. Additionally, the electron beam causes a negative charge to build up on the surface of most specimens, resulting in lower image resolution. To overcome this problem, specimens are routinely coated with a thin layer of metal to permit them to conduct away the negative charge. These metal coatings usually harm wet and living specimens and further imped the electron micro-imaging of such substances. Consequently, microscopic examination of wet or living substances has generally been performed with traditional optical light microscopes which lack the needed magnification and resolution power of electron microscopes.
The problem of developing a high resolution imaging capability for living and wet substances or for surface morphology of materials has preoccupied the field of micro-imaging technology for some time. Attempts to solve the problem have led to the creation of such devices as acoustic microscopes, electron tunneling microscopes, heavy ion microscopes and x-ray microscopes. However, these devices are not capable of producing high resolution images without subjecting specimens to high vacuum or heavy radiation doses and usually both.